5-cnac as oral delivery agent for parathyroid hormone fragments

ABSTRACT

Pharmaceutical compositions for the effective oral delivery of a parathyroid hormone, PTH, as well as methods for administration of the compositions are provided. Additionally, methods for stimulating new bone formation and treating and/or preventing osteoporosis are also provided.

BACKGROUND OF THE INVENTION

[0001] 1. Field of the Invention

[0002] The present invention relates to the oral delivery of parathyroidhormone (PTH). The mammalian parathyroid hormones, e.g. human (hPTH),bovine (bPTH) and porcine (pPTH), are single polypeptide chains of 84amino acid residues having molecular weights of approximately 9500.Specifically, the present invention relates to PTH fragmentsincorporating at least the first 28 N-terminal amino acid residues (PTH(1-28)) up to and including the first 41 N-terminal amino acid residues(PTH (1-41)). More particularly, the invention is directed topharmaceutical compositions for the oral delivery of PTH, saidcompositions comprising PTH (1-28) to (1-41) andN-(5-chlorosalicyloyl)-8-aminocaprylic acid (5-CNAC).

[0003] 2. Description of the Related Art

[0004] PTH studies done in animals and humans with PTH, PTH-relatedpeptides, and PTH analogues have demonstrated its usefulness inincreasing bone formation and bone resorption and have prompted interestin its use for the treatment of osteoporosis and related bone disorders.However, the oral delivery of PTH in mammals has proven difficult due,at least in part, to the insufficient stability of PTH in thegastrointestinal tract as well as the inability of PTH to be readilytransported through the intestinal walls into the blood stream.

[0005] U.S. Pat. No. 5,773,647 (the '647 patent) describes 193 carriercompounds useful for the delivery of active agents, including PTH. Oneof the carrier compounds expressly described therein isN-(5-chlorosalicyloyl)-8-aminocaprylic acid (5-CNAC) having the formula

[0006] Example 2 in Column 6 of the '647 patent describes thepreparation of 11 different dosing compositions some intracolonic (IC)and some oral gavage (PO) each containing parathyroid hormone and acarrier, the carrier being different for each composition. An IC dosingcomposition was prepared using 5-CNAC as the carrier. Example 3 thereindescribes in vivo tests carried out dosing male Sprague-Dawley rats withthe dosing solutions prepared in Example 2. Blood samples were collectedand the serum PTH concentration was quantified for each rat.

[0007] Surprisingly, it has now been found that 5-CNAC in combinationwith specific PTH fragments, i.e. PTH fragments incorporating at leastthe first 28 N-terminal amino acid residues (PTH (1-28)) up to andincluding the first 41 N-terminal amino acid residues (PTH (1-41)) whenorally administered gives unexpectedly high PTH serum levels relative toother PTHs and other carriers and provide a sharp C_(max) allowing for abone formation effect.

SUMMARY OF THE INVENTION

[0008] Accordingly, the present invention is directed to pharmaceuticalcompositions suitable for oral delivery of PTH fragments and to methodsof administering such compositions.

[0009] Specifically, the instant invention is directed to apharmaceutical composition for oral delivery comprising atherapeutically effective amount of a PTH fragment and 5-CNAC, said PTHfragment selected from PTH (1-28) to PTH (1-41). Preferably, the PTH ishuman parathyroid hormone, hPTH.

[0010] In another embodiment, the invention is directed to a method fororally administering an effective dose of PTH comprising orallyadministering to a patient in need of PTH a pharmaceutical compositioncomprising a therapeutically effective amount of a PTH fragment and5-CNAC, said PTH fragment selected from PTH (1-28) to PTH (1-41).

[0011] The invention is also directed to a method of stimulating newbone formation comprising orally administering to a patient in need ofnew bone formation a pharmaceutical composition comprising atherapeutically effective amount of a PTH fragment and 5-CNAC, said PTHfragment selected from PTH (1-28) to PTH (1-41).

[0012] In a further embodiment, the invention is directed to a method oftreatment or prevention of osteoporosis comprising orally administeringto a patient in need of said treatment or prevention a pharmaceuticalcomposition comprising a therapeutically effective amount of a PTHfragment and 5-CNAC, said PTH fragment selected from PTH (1-28) to PTH(1-41).

[0013] In a still further embodiment, the invention is directed to theuse of 5-CNAC for the preparation of a pharmaceutical compositionsuitable for the oral delivery of PTH fragments selected from PTH (1-28)to PTH (1-41).

[0014] Further features and advantages of the invention will becomeapparent from the following detailed description of the invention.

DETAILED DESCRIPTION OF THE INVENTION

[0015] The PTH fragments can be of any parathyroid hormone, particularlymammalian parathyroid hormone, e.g. human (hPTH), bovine (bPTH), andporcine (pPTH) and particularly hPTH and will incorporate at least thefirst 28 N-terminal residues (PTH (1-28)) up to and including the first41 N-terminal residues (PTH (1-41)) and include without limitation PTH(1-28), PTH (1-31), PTH (1-34), PTH (1-37), PTH (1-38) and PTH (1-41).Human parathyroid hormone (1-34) is particularly preferred. Theseparathyroid hormone fragments are commercially available or can beobtained recombinantly or by peptide synthesis.

[0016] For purposes of the instant invention, the 5-CNAC, i.e.N-(5-chlorosalicyloyl)-8-aminocaprylic acid, can be the free acid,analogs thereof, its monosodium and disodium salts, ethanol solvates ofthe sodium salts and the monohydrates of the sodium salts and anycombinations thereof. The free acid, the disodium salt of 5-CNAC and themonohydrate thereof are particularly useful.N-(5-chlorosalicyloyl)-8-aminocaprylic acid is described in theaforementioned '647 patent, the contents of which are herebyincorporated by reference, and can be made by methods described therein.The sodium salts and alcohol solvates and hydrates thereof are describedin WO 00/059863, along with methods for preparing them.

[0017] The disodium salt may be prepared from the ethanol solvate byevaporating or drying the ethanol solvate by methods known in the art toform the anhydrous disodium salt. Drying is generally carried out at atemperature of from about 80 to about 120° C., preferably from about 85to about 90° C., and most preferably at about 85° C. The drying step isgenerally performed at a pressure of 26″ Hg or greater. The anhydrousdisodium salt generally contains less than about 5% by weight of ethanoland preferably less than about 2% by weight of ethanol, based on 100%total weight of anhydrous disodium salt.

[0018] The disodium salt of N-(5-chlorosalicyloyl)-8-aminocaprylic acidcan also be prepared by making a slurry ofN-(5-chlorosalicyloyl)-8-aminocaprylic acid in water and adding twomolar equivalents of aqueous sodium hydroxide, sodium alkoxide or thelike. Suitable sodium alkoxides include, but are not limited to, sodiummethoxide, sodium ethoxide, and combinations thereof.

[0019] A still further method of preparing the disodium salt is byreacting N-(5-chlorosalicyloyl)-8-aminocaprylic acid with one molarequivalent of sodium hydroxide to form a monosodium salt and then addingan additional one molar equivalent of sodium hydroxide to yield thedisodium salt.

[0020] The disodium salt can be isolated as a solid by concentrating thesolution containing the disodium salt to a thick paste by vacuumdistillation. This paste may be dried in a vacuum oven to obtain thedisodium salt of N-(5-chlorosalicyloyl)-8-aminocaprylic acid as a solid.The solid can also be isolated by spray drying an aqueous solution ofthe disodium salt.

[0021] The ethanol solvates, as described in the aforementioned WO00/059863, include, but are not limited to, a molecular or ionic complexof molecules or ions of ethanol solvent with molecules or ions of thedisodium salt of N-(5-chlorosalicyloyl)-8-aminocaprylic acid. Typically,the ethanol solvate contains about one ethanol molecule or ion for everymolecule of disodium salt of N-(5-chlorosalicyloyl)-8-aminocaprylicacid.

[0022] The ethanol solvate of the disodium salt ofN-(5-chlorosalicyloyl)-8-aminocaprylic acid can be prepared bydissolving N-(5-chlorosalicyloyl)-8-aminocaprylic acid in ethanol.Typically, each gram of N-(5-chlorosalicyloyl)-8-aminocaprylic acid isdissolved in from about 1 to about 50 mL of ethanol and generally, fromabout 2 to about 10 mL of ethanol. TheN-(5-chlorosalicyloyl)-8-aminocaprylic acid/ethanol solution is thenreacted with a molar excess of a sodium containing salt, such as amonosodium containing salt, relative toN-(5-chlorosalicyloyl)-8-aminocaprylic acid, i.e. for every mole ofN-(5-chlorosalicyloyl)-8-aminocaprylic acid there is more than one moleof sodium cations, yielding the ethanol solvate. Suitable monosodiumsalts include, but are not limited to, sodium hydroxide; sodiumalkoxides, such as sodium methoxide and sodium ethoxide; and anycombination of the foregoing. Preferably, at least about two molarequivalents of the monosodium containing salt are added to the ethanolsolution, i.e. for every mole of N-(5-chlorosalicyloyl)-8-aminocaprylicacid there is at least about two moles of sodium cations. Generally, thereaction is performed at or below the reflux temperature of the mixture,such as at ambient temperature. The ethanol solvate is then recovered bymethods known is the art, such as, concentration of the resulting slurryat atmospheric distillation, cooling the concentrated slurry andfiltering the solid. The recovered solid can then be vacuum dried toobtain the ethanol solvate.

[0023] The hydrates of the disodium salts of theN-(5-chlorosalicyloyl)-8-aminocaprylic acid may be prepared by dryingthe ethanol solvate to form an anhydrous disodium salt, as describedabove, and hydrating the anhydrous disodium salt. Preferably, themonohydrate of the disodium salt is formed. Since the anhydrous disodiumsalt is very hydroscopic, the hydrate forms upon exposure to atmosphericmoisture. Generally, the hydrating step is performed at from aboutambient temperature to about 50° C., preferably ambient temperature toabout 30° C. and in an environment having at least 50% relativehumidity. Alternatively, the anhydrous disodium salt may be hydratedwith steam.

[0024] The amount of PTH fragment to be administered is generally anamount effective to stimulate new bone formation i.e. a therapeuticallyeffective amount. This amount will necessarily vary with the age, size,sex and condition of the subject to be treated, the nature and severityof the disorder to be treated and the like. However, the amount can beless than that amount when a plurality of the compositions are to beadministered, i.e., the total effective amount can be administered incumulative dosage units. The amount of PTH can also be more than theeffective amount when the composition provides sustained release of thepharmacologically active agent The total amount of PTH to be used can bedetermined by methods known to those skilled in the art. However, ingeneral, satisfactory results will be obtained systemically at dailydosages of from about 0.001 μg/kg to about 10 mg/kg animal body weight,preferably 1 μg/kg to about 6 μg/kg body weight.

[0025] The pharmaceutical compositions of the present inventiontypically contain a delivery effective amount of 5-CNAC, i.e. an amountsufficient to deliver the PTH for the desired effect Generally, the5-CNAC is present in an amount of 2.5% to 99.4% by weight, morepreferably 25% to 50% by weight of the total composition.

[0026] Oral administration of the pharmaceutical compositions accordingto the invention can be accomplished regularly, e.g. once or more on adaily or weekly basis; intermittently, e.g. irregularly during a day orweek; or cyclically, e.g. regularly for a period of days or weeksfollowed by a period without administration.

[0027] The dosage form of the pharmaceutical compositions of the instantinvention can be any known form, e.g. liquid or solid dosage forms.

[0028] The liquid dosage forms include solution emulsions, suspensions,syrups and elixirs. In addition to the PTH and 5-CNAC, the liquidformulations may also include inert excipients commonly used in the artsuch as, solubilizing agents e.g. ethanol; oils such as cottonseed,castor and sesame oils; wetting agents; emulsifying agents; suspendingagents; sweeteners; flavorings; and solvents such as water.

[0029] The solid dosage forms include capsules, soft-gel capsules,tablets, caplets, powders, granules or other solid oral dosage forms,all of which can be prepared by methods well known in the art.

[0030] The pharmaceutical compositions may additionally compriseadditives in amounts customarily employed including, but not limited to,a pH adjuster, a preservative, a flavorant, a taste-masking agent, afragrance, a humectant, a tonicifier, a colorant, a surfactant, aplasticizer, a lubricant such as magnesium stearate, a flow aid, acompression aid, a solubilizer, an excipient, a diluent such asmicrocrystalline cellulose, e.g. Avicel PH 102 supplied by FMCcorporation, or any combination thereof. Other additives may includephosphate buffer salts, citric acid, glycols, and other dispersingagents.

[0031] The composition may also include one or more enzyme Inhibitors,such as actinonin or epiactinonin and derivatives thereof; aprotinin,Trasylol and Bowman-Birk inhibitor.

[0032] Further, a transport inhibitor, i.e. a p-glycoprotein such asKetoprofin, may be present in the compositions of the present invention.

[0033] The solid pharmaceutical compositions of the instant inventioncan be prepared by conventional methods e.g. by blending a mixture ofthe PTH fragment, the 5-CNAC, and any other ingredients, kneading, andfilling into capsules or, instead of filling into capsules, moldingfollowed by further tableting or compression-molding to give tablets. Inaddition, a solid dispersion may be formed by known methods followed byfurther processing to form a tablet or capsule.

[0034] Preferably, the ingredients in the pharmaceutical compositions ofthe instant invention are homogeneously or uniformly mixed throughoutthe solid dosage form.

[0035] Parathyroid hormones are indicated for preventing or treating allbone conditions which are associated with increased calcium depletion orresorption or in which stimulation of bone formation and calciumfixation in the bone is desirable, e.g. osteoporosis of various genesis(e.g. juvenile, menopausal, post-menopausal, post-traumatic, caused byold age or by corticoid-steroid therapy or inactivity), fractures,osteopathy, including acute and chronic states associated with skeletaldemineralization, osteo-malacia, periodontal bone loss or bone loss dueto arthritis or osteoarthritis or cancer (e.g. bone metastis) or fortreating hypoparathyroidism.

[0036] Parathyroid hormones are particularly indicated for preventing ortreating osteoporosis of various genesis.

[0037] According to a further embodiment of the invention, the PTH maybe employed as adjunct or adjuvant to other therapy, e.g. a therapyusing a bone resorption inhibitor, for example as in osteoporosistherapy, in particular a therapy employing calcium, a calcitonin or ananalogue or derivative thereof, e.g. salmon, eel or human calcitonin, asteroid hormone, e.g. an estrogen, a partial estrogen agonist orestrogen-gestagen combination, a SERM (Selective Estrogen ReceptorModulator) e.g. raloxifene, lasofoxifene, TSE-424, FC1271, Tibolone(Livial®), vitamin D or an analogue thereof or an activator of PTHrelease, or bisphosphonates, e.g. clodronic acid, etidronic acid,pamidronic acid, aledronic acid, ibandronic acid, zoledronic acid,risedronic acid or tiludronic acid and salts and hydrates thereof.

[0038] When the PTH is administered in conjunction with, e.g. as anadjuvant to bone resorption inhibition therapy, dosages for theco-administered inhibitor will of course vary depending on the type ofinhibitor drug employed, e.g. whether it is a steroid or a calcitonin,on the condition to be treated, whether it is a curative or preventivetherapy, on the regimen and so forth.

[0039] The oral administration of the present invention may be to anyanimal in need thereof, including, but not limited to, mammals, such asrodents, cows, pigs, dogs, cats, and primates, particularly humans.

[0040] The following examples serve to further illustrate the invention.

EXAMPLE 1

[0041] The following capsules are prepared as follows:

[0042] Capsules prepared from 800 μg hPTH* (Capsule 1A)

[0043] Capsules prepared from 400 mg 5-CNAC**/800 μg hPTH* (Capsule 1B)

[0044] The hPTH only capsules are prepared by weighing out 400 μg of thehPTH and placing it directly into each capsule. The hPTH/5-CNAC capsulesare prepared as dry blends by weighing out the individual componentsblending them together to make a homogeneous mix and then hand filling400 mg of the mix into each capsule;

EXAMPLE 2 Primate Administration

[0045] The capsules prepared in Example 1 are administered to Rhesusmonkeys as follows: four monkeys in a group are each dosed with onecapsule prepared as in Example 1 as follows:

[0046] The Rhesus monkeys fast overnight prior to dosing and arerestrained in chairs fully conscious, for the duration of the studyperiod. The capsules are orally administered via a gavage tube followedby 10 mL of water.

[0047] Blood samples are collected at 0, 0.25, 0.5, 0.75, 1, 1.5, 2, 3,4, 5, and 6 hours after administration. Plasma hPTH levels aredetermined by radioimmunoassay. The primate plasma PTH results from eachgroup of monkeys are averaged and the maximum mean plasma calculated.The results for the PTH only group are reported in Table 1 and theresults for the hPTH/5-CNAC group are reported in Table 2. TABLE 1 hPTHONLY hPTH PLASMA CONCENTRATIONS (pg/mL) AFTER ORAL ADMINISTRATION TO THERHESUS MONKEY Dose: 1 Capsule 1 D Animal Time [hours] no. 0 0.25 0.500.75 1 1.5 2 3 4 5 6 R927 0 0 0 0 0 0 0 0 0 0 0 S982 0 0 0 0 0 0 0 0 0 00 Mean 0 0 0 0 0 0 0 0 0 0 0 SD 0 0 0 0 0 0 0 0 0 0 0 SEM 0 0 0 0 0 0 00 0 0 0

[0048] TABLE 2 hPTH / 5-CNAC hPTH PLASMA CONCENTRATIONS (pg/mL) AFTERORAL ADMINISTRATION TO THE RHESUS MONKEY Dose: 1 Capsule 1 B Ani- malTime [hours] no. 0 0.25 0.50 0.75 1 1.5 2 3 4 5 6 R944 0 83 191 300 360262 154 35 0 0 0 S963 0 127 332 663 1258 150 34 0 0 0 0 Mean 0 105 262482 809 206 94 17 0 0 0 SD 0 31 100 257 635 79 85 25 0 0 0 SEM 0 22 71182 449 56 60 17 0 0 0

[0049] As can be seen from the data in Tables 1 and 2, the 5-CNACsignificantly facilitates the oral delivery of the hPTH fragment. Inaddition, the data in Table 2 indicate a sharp C_(max) in the PTH plasmaprofile allowing for a bone formation effect.

[0050] The foregoing embodiments and examples are given merely toillustrate the instant invention and are not intended to be limiting.Numerous other embodiments and variations are within the scope of theinvention and readily accessible to those skilled in the art.

We claim:
 1. A pharmaceutical composition for oral delivery comprising atherapeutically effective amount of a PTH fragment and 5-CNAC, said PTHfragment selected from PTH (1-28) to PTH (1-41).
 2. A pharmaceuticalcomposition according to claim 1 wherein the PTH is selected from PTH(1-28), PTH (1-31), PTH (1-34), PTH (1-37), PTH (1-38) and PTH (1-41).3. A pharmaceutical composition according to claim 1 wherein the PTH isPTH (1-34).
 4. A pharmaceutical composition according to claim 1 whereinthe PTH is recombinant PTH.
 5. A pharmaceutical composition according toclaim 3 wherein the PTH is recombinant PTH.
 6. A pharmaceuticalcomposition according to claim 1 wherein the PTH is human parathyroidhormone.
 7. A pharmaceutical composition according to claim 6 whereinthe human parathyroid hormone is hPTH (1-34).
 8. A pharmaceuticalcomposition according to claim 1 wherein the 5-CNAC is selected from thefree acid, the disodium salt of N-(5-chlorosalicyloyl)-8-aminocaprylicacid and the monohydrate thereof.
 9. A pharmaceutical compositionaccording to claim 1 wherein the 5-CNAC isN-(5-chlorosalicyloyl)-8-aminocaprylic acid.
 10. A pharmaceuticalcomposition according to claim 1 wherein the 5-CNAC is the disodium saltof N-(5-chloro-salicyloyl)-8-aminocaprylic acid.
 11. A method for orallyadministering an effective dose of PTH comprising orally administeringto a patient in need of PTH a pharmaceutical composition comprising atherapeutically effective amount of a PTH fragment and 5-CNAC, said PTHfragment selected from PTH (1-28)-PTH (1-41).
 12. A method according toclaim 11 wherein the PTH is selected from PTH (1-28), PTH (1-31), PTH(1-34), PTH (1-37), PTH (1-38) and PTH (1-41).
 13. A method according toclaim 11 wherein the PTH is hPTH (1-34).
 14. A method according to claim11 wherein the PTH is recombinant PTH.
 15. A method according to claim11 wherein the PTH is recombinant hPTH.
 16. A method according to claim11 wherein the 5-CNAC is selected from the free acid, the disodium saltof N-(5-chlorosalicyloyl)-8-aminocaprylic acid and the monohydratethereof.
 17. A method according to claim 11 wherein the 5-CNAC isN-(5-chloro-salicyloyl)-8-aminocaprylic acid.
 18. A method according toclaim 11 wherein the 5-CNAC Is the disodium salt ofN-(5-chlorosalicyloyl)-8-aminocaprylic acid.
 19. A method of stimulatingnew bone formation comprising orally administering to a patient in needof new bone formation a pharmaceutical composition comprising atherapeutically effective amount of a PTH fragment and 5-CNAC, said PTHfragment selected from PTH (1-28)-PTH (1-41).
 20. A method of treatmentor prevention of osteoporosis comprising orally administering to apatient in need of said treatment or prevention a pharmaceuticalcomposition comprising a therapeutically effective amount of a PTHfragment and 5-CNAC, said PTH fragment selected from PTH (1-28)-PTH(1-41).
 21. The use of 5-CNAC for the preparation of a pharmaceuticalcomposition suitable for the oral delivery of PTH fragments selectedfrom PTH (1-28)-PTH (1-41).